The following description of the background of the invention is provided to aid in understanding the invention, but is not admitted to be or to describe prior art to the invention.
Cellular signal transduction is a fundamental mechanism whereby extracellular stimuli are relayed to the interior of cells. One of the key biochemical mechanisms of signal transduction involves the reversible phosphorylation of proteins. Phosphorylation of amino acids regulates the activity of mature proteins by altering their structure and function.
Phosphate most often resides on the hydroxyl moiety of serine, threonine, or tyrosine amino acids in proteins. Enzymes that mediate phosphorylation of cellular effectors fall into two classes. While protein phosphatases hydrolyze phosphate moieties from phosphoryl protein substrates, protein kinases transfer a phosphate moiety from adenosine triphosphate to protein substrates. The converse functions of protein kinases and protein phosphatases balance and regulate the flow of signals in signal transduction processes.
Protein kinases are divided further into two groups: receptor and non-receptor type proteins. Receptor protein kinases comprise an extracellular region, a transmembrane region, and an intracellular region. Part of the intracellular regions of receptor protein kinases harbor a catalytic domain.
Receptor protein kinases are also divided into three classes based upon the amino acids they act upon. Some phosphorylate serine or threonine only, some phosphorylate tyrosine only, and some phosphorylate serine, threonine, and tyrosine.
Receptor protein tyrosine kinases (RPTKs) are typically activated in the cell when a ligand binds to the extracellular region of the receptor. A model for ligand mediated activation of RPTKs features the ligand bringing the receptors within close proximity to one another. Some ligands are dimers and thereby bring the receptors that bind them into close proximity with one another. By bringing two RPTKs together, ligands place RPTK intracellular catalytic regions in close proximity to one another such that they cross-phosphorylate. Cross phosphorylation requires not only the dimerization process but also the occurrence of a conformational change preceding phosphorylation. The necessity of the conformational change preceding phosphorylation is illustrated by the fact that some RPTKs, such as the insulin receptor, are pre-dimerized and inactive before binding their activating ligands.
The presence of phosphate moieties on the RPTK intracellular regions constitutes a cellular signal that causes other signal transduction molecules to bind to the RPTK. In this manner, RPTKs propagate the extracellular signal to the cell nucleus, thereby generating messages encoding proteins that cause cellular effects.
Because RPTKs control a variety of cellular function, any alteration in the normal function of an RPTK can result in an abnormal condition in an organism. For example, differentiation and survival of neuronal cells is dependent upon the proper function of RPTKs. Specifically, it has been shown that the interaction between activated Trk (Barbacid et al., Biochimica et Biophysica Acta, 1072:115-127, 1991) and a signaling component such as SHC is important for promoting neuronal cell differentiation and survival. Transgenic mice containing knockout mutations in genes encoding each of the known Trk receptors or the Trk ligand displayed severe neurological dysfunction and, in all cases, important types of neural tissue were absent. Smeyne et al., 1994, Nature 368: 246-249; Klein et al., 1994, Nature 268: 249-251, 1994.
Specific neurotrophic factors (generally referred to as ligands herein) have also been shown to promote neuronal survival. In particular, glial-derived neurotrophic factor (GDNF) has been identified as a neuronal survival factor. Jing et al., 1996, Cell 85:1113-1124; Trupp et al., 1996, Nature 381:785-789; Durbec et al., 1996, Nature 381: 789-793. GDNF has been shown to bind to a complex of C-RET and another cell surface protein GDNFR-.alpha., which has no intracellular domain.
In an effort to discover novel treatments for diseases, biomedical researchers have designed, synthesized, and tested molecules that modulate protein kinase function in cells. Some organic molecules have been identified as modulators of RPTK function. For example, bis monocyclic, bicyclic, or heterocyclic aryl compounds (PCT WO 92/20642), vinylene-azaindole derivatives (PCT WO 94/14808), 1-cyclopropyl-4-pyridylquinolones (U.S. Pat. No. 5,330,992), styryl compounds (U.S. Pat. No. 5,217,999), styryl-substituted pyridyl compounds (U.S. Pat. No. 5,302,606), certain quinazoline derivatives (EP Application No. 0 566 266 Al), seleoindoles and selenides (PCT WO 94/03427), tricyclic polyhydroxylic compounds (PCT WO 92/21660), and benzylphosphonic acid compounds (PCT WO 91/15495) have been identified as compounds which modulate the function of protein kinases.
There remains a great need in the medical field for identifying compounds that modulate the function of RPTKs regulating neuronal survival, in particular. Many patients suffer from diseases caused by neuronal degeneration, such as Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis, for example. Although neurotrophic factors have been proposed as therapeutic agents for neuronal diseases, they often cannot reach their target receptors since they rapidly degrade in the blood stream and cannot pass through cell membranes or the blood brain barrier. Thus, identifying effective therapeutics for treating neuronal diseases lies in identifying non-peptide compounds which do not rapidly degrade in the blood stream and which can pass through cell membranes and the blood-brain barrier.